Abstract
The intercellular communication mediated by extracellular vesicles (EVs) has gained international interest during the last decade. Interfering with the mechanisms regulating this cellular process might find application particularly in oncology where cancer cell‐derived EVs play a role in tumour microenvironment transformation. Although several mechanisms were ascribed to explain the internalization of EVs, little is our knowledge about the fate of their cargos, which are crucial to mediate their function. We recently demonstrated a new intracellular pathway in which a fraction of endocytosed EV‐associated proteins is transported into the nucleoplasm of the host cell via a subpopulation of late endosomes penetrating into the nucleoplasmic reticulum. Silencing tetraspanin CD9 both in EVs and recipient cells strongly decreased the endocytosis of EVs and abolished the nuclear transfer of their cargos. Here, we investigated whether monovalent Fab fragments derived from 5H9 anti‐CD9 monoclonal antibody (referred hereafter as CD9 Fab) interfered with these cellular processes. To monitor the intracellular transport of proteins, we used fluorescent EVs containing CD9‐green fluorescent protein fusion protein and various melanoma cell lines and bone marrow‐derived mesenchymal stromal cells as recipient cells. Interestingly, CD9 Fab considerably reduced EV uptake and the nuclear transfer of their proteins in all examined cells. In contrast, the divalent CD9 antibody stimulated both events. By impeding intercellular communication in the tumour microenvironment, CD9 Fab‐mediated inhibition of EV uptake, combined with direct targeting of cancerous cells could lead to the development of novel anti‐melanoma therapeutic strategies.
Highlights
Growing evidence indicate that intercellular communication in multicellular organisms is mediated by direct cell‐cell contact or soluble molecules, and by extracellular vesicles (EVs), ie lipid bilayer‐enclosed nanobiological units actively re‐ leased from all cell types.[1,2] In contrast to soluble signalling mol‐ ecules, bioactive compounds associated with EVs are protected from degradation.[3,4] EVs are found in internal and external bodily fluids and act as mediators of long‐distance transfer of biological information
We previously reported that cargo proteins derived from EVs are internalized by host cells, and a fraction of them is transferred to their nucleoplasm by the intermediate of late en‐ dosomes entering into nucleoplasmic reticulum.[23,29]
When the same experiments were performed with primary mesenchymal stromal cells (MSCs) as recipient cells as well as donor cells for fluorescent EVs (1 × 109 particle per mL), we observed a significant decrease in nuclear and cytoplasmic CD9‐GFP in cells pre‐treated with CD9 Fab (25 μg/mL) (Figure 3D; data not shown)
Summary
By monitoring the internalization of melanoma‐derived EVs and the intracellular routes of their content, CD9 (see below), we discovered that EV‐associated proteins are transported into the nucleus of the host cell through late endosomes entering the nucleoplasmic reticulum (Figure 1A).[23] Therein, EV‐associated cargo molecules can modify the gene expression of the host cells These surprising findings are in line with numerous studies showing the atypical nuclear localiza‐ tion of the EV‐associated proteins CD9 and CD133 as well as the shuttling of proteins and nucleic acids to nucleoplasm of recipient cells.[3,24,25,26,27,28] Recently, we described that two proteins, ie vesicle‐as‐ sociated membrane protein‐associated protein A (VAP‐A) and the cytoplasmic oxysterol‐binding protein‐related protein 3 (ORP3), are essential for the entry and the tethering of late endosomes to nuclear envelope invaginations of type II (Figure 1B). | 4410 new modality in cancer treatment by inhibiting the intercellular communication within the cancer cell niche
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
